Project Summary Glaucoma is a major cause of blindness, characterized by the death of retinal ganglion cells (RGCs) and loss of vision. An elevated intraocular pressure (IOP) is the biggest risk factor in the disease, but localized inflammatory signaling can also contribute to the pathology. This proposal will investigate the pathways linking elevated IOP to local inflammatory signaling and RGC death, based on the innovative hypothesis that ATP-dependent release of the inflammatory cytokine ?IL-1? from microglial cells links mechanical strain to vision loss in glaucoma. Stretch- dependent ATP release from optic nerve head astrocytes will be compared using astrocytes missing genes implicated in mechanosensitive ATP release. The most promising candidates will be examined in vivo using a mouse model for chronic ocular hypertension; reduced levels of inflammatory responses or RGC death will implicate mechanosensitive ATP release. The ability of ATP to enhance microglial migration and IL-1? release will be explored in isolated retinal microglial cells to enable detailed characterization. The contribution to RGCs death in a model of chronic ocular hypertension will be confirmed. The ability of IL-1? to kill RGCs in vivo will be confirmed and its contribution to death in ocular hypertension tested. The effect of substrate stiffness on the magnitude of mechanosensitive ATP release will be investigated in vitro and in vivo. Finally, the links between ATP release and IL-1? will be confirmed in the retina of glaucomatous humans. In summary, this proposal will connect elevated IOP with proinflammatory responses through aberrant purinergic signaling. This novel hypothesis will advance our understanding of glaucoma while identifying possible new targets for intervention.